Innate Immunity - Science topic

According to previous articles in Science or Nature, there are innate cells that may have "memory" phenotype. These cells include both so called educated NK lymphocytes as well as some myeloid cells such as monocytes. However this memory is not the same and does not last for years as in the case of adaptive lymphocytes. One of the reason is fact that innate cells do not undergo clonal expansion.

The innate immune system serves as the body's initial defense against a wide range of pathogens, including bacteria, viruses, and fungi. It operates through a combination of physical barriers, chemical defenses, and cellular responses:

Overall, the innate immune system provides rapid, nonspecific defense mechanisms to protect the body from various pathogens, initiating the immune response until the adaptive immune system can mount a more specific defense.

Hello Robbie Manuel

T helper 17 cells (Th17) are the most affected. AhR activation may directly or indirectly also modulate the commitment of Tregs, Th1 and Th2.

Please refer to the articles attached for more information.

Best.

Also, have a look at this useful link.

Dear all,

I haven't found a paper yet that claims THP1 is a classical monocyte. Would you know of one? Thank you!

you can have a look on :

Good Question...

You can try Ischaemia re-perfusion injury model.

So...yes and no... in immunology germline is often used to describe origin of the immune population. So, germline immune cells are those that do not arise out of the bone marrow, and they usually are tissues resident cells, like alveolar macrophages, langerhan cells, etc.

So the PRR technically is germline encoded since they don't change, and it is passed down...which is why almost everyone respond to the same pathogen motifs.

They seem to be common monocyte-macrophage progenitors (CD11b neg Ly6C+). I observed this population of cells as well in the spleen of mice.

Here a link to a book which explains that.

Best,

Hi Vivek,

Sorry for my late answer. I am in the same case as you; i did not get any answer from them. Is it working your differentiation from CLP ?

Thank you very much !

Julie

Szczepan, thank you for replying to the question. Do you have any reading material that supports your answer? I will highly appreciate if you could share here. Thank you again.

Thank you very much Ricardo for your answer. Does it means that wild plants living in the absence of challenges will gradually lose their ability to fight against diseases and pest? could it be an epigenetic phenomenon, like the methylation of CpGs of the promoters of resistance genes?

This depends on the type of immune cells you expect. I would look at IL-8 and MPO for neutrophils. ECP, EDN, IL-5 for eosinophils, Granzyme B for e.g. NK cells, histamine for basophils and mast cells. To sample tears you can use schirmer strips and then you can extract the analytes.

Check this paper: https://www.physiology.org/doi/abs/10.1152/ajpgi.1997.273.2.G322?url_ver=Z39.88-2003&rfr_id=ori%3Arid%3Acrossref.org&rfr_dat=cr_pub%3Dpubmed

Hi Thomas,

I have acquired MC/9 cells from ATCC and, unfortunately, according to selling contidions, I am not allow to distribute theses cells. I am sorry.

Geraldo.

this link is useful

https://www.ncbi.nlm.nih.gov/pubmed/24973890

regards

STAT 3 pathways

Hi Dr. Al-Salhie, unfortunately your link shows an equipment that is called "Crocodile" ( the commercial name of the instrument is crocodile, a device to pipette, wash and incubate plates only). However, Berthold technologies does not sell ELISA assays directed to crocodilian proteins, just as nobody else does for reptiles. Believe me, I searched everywhere for antibodies against snake proteins.

Thank you though, for a split second my hopes were high...

Chemistry World has reported on this issue:

and I have a follow up at my blog as one of the two references cited in support of the SmartFlares includes plagiarism... of my blog.

1. What you found is expected. I am sharing my IL-10 experience with human macrophages. Human macrophages are very different from murine macrophages. IL-10 is a murine M2 marker. It does not work in human. M2s have subtypes: M2a (IL-4), M2b (LPS+Immune complex), M2c(IL-10/TGF-b). I saw, IL-4-treated human M2s dnt express IL-10. But if unpolarized macs are stimulated with IL-10, they produce IL-10. 

2. PMA-treated macs are similar to monocyte derived macs (MDMs), but not the same. Surface marker expression, cytokine release of PMA-macs are different from MDMs. 

3. If you do qPCR, I think you need unpolarized cells as negative control/ basal expression. 

You can check this article: PMID: 25870903

Simplified suggested protocol with not so many steps.

1. Mix the standard isolated neutrophil cell suspension of fish with stimulant at a ratio 1:4. Total volume 50 microliter.  Use : 1 μg/mL of PMA final concentration as a stimulant. Both the cell suspension and PMA should have been diluted and kept in HBSS with Ca, Mg, without phenol red.

2. Shake  for 10 seconds on a shaker.

3. Incubate the suspension at room temperature for 2 hours.

4. Your NETS are now ready to be stained with whichever dye you are using and whichever protocol you are using for cell fixation.

P.S. instead of fish blood, its better to use fish anterior kidneys to isolate neutrophils.

Hi Kerolous 

wild type virus usually is more immunogen  than culturally adapted

Thank you!

Have you seen this paper yet?

You may consider using such agent in a kidney disease experimental model.

Hi Wolfgang,

Have you considered STAT phosphorylation as read out? We are doing lots of cytokine-induced phospho-flow assays in our lab, albeit using primary human T cells mostly. We haven't tried out IFNg but type 1 IFNs result in robust STAT1 phosphorylation within minutes and is easily detectable by flow cytometry. Similarly, IL-6 induces pSTAT3 very potently in T cells (and monocytes). However, I would not think that DCs respond well to 'normal' (classical) IL-6 signaling. Although activated DCs produce a lot of IL-6, they do not express (much) of the IL-6 receptor (IL-6R) on their surface. They are, however, susceptible to IL-6 'transsignaling' using a complex of IL-6 with soluble IL-6R...still, to test your JAK inhibitors, I would probably go for type I INF (via JAK1/TYK2) or IFNg (via JAK1/JAK2) induced pSTAT1.

Hope this is useful.

Christian

Can you, for example, look, if LPS or PMA can activate ROS - production by macrophages?

The RAW 264.7 cell line is one of the best characterized cell lines representing the M1 phenotype. For the M2 phenotype, the IC-21the cell line is a good representative line(Yagnik, Landis. Arthritis Rheum. 2000 Aug;43(8):1779-89)

Hi Dear,

Please read the attached file, I think it will be helpful.

I have one more question Mario. Do know any paper regarding these organism that can be helpful to me. I really appreciate if you tell me the name of the paper; I probably has not seen it or you probably have some clue about a few papers regarding this . 

Thanks,

Dear,Naael Ali

There are many different types of auto-antigens in health individual which are under suppressor manner, that when uncovered may lead to production of immune response( immune reaction ) and presence of different types of auto-antibodies (because these auto- antigens are hidden or occult  antigens with latent  bioactive functioning ). For examples, sperms (components: antisperm antibody), joints synovial fluid, ovum (components) and ocular fluids when this compartments are being (conditions) associated with trauma, infections,rupture, abrasions or any pathological causes that may lead the (body) immune system to known those hidden or covered antigens and reflect to induce an auto-immunological reactions.

Good, Luck

NASEER ALMUKHTAR 

Thanks for the additional comments! Unfortunately I don't think these approaches would be specific enough, because I'm talking about tissue sections and the mentioned downstream signaling molecules are not exclusively used by TLRs, so their activation doesn't prove TLR activation.

I have tried transfection of cellular RNA into 293T cells by lipofectamine, the cellular RNA did not stimulate the cells to produce IFN. the transfection of RNA by lipofectamine may directly deliver the RNA into cytoplasm that avoid to be recognized by TLRs. 

Hi Awatif,

This paper is very helpful. The only pity is that effect of IL12 or IL15 on pDC maturation is not evaluated there.

QM

Do you guys know how many Nk cells we can get per spleen using the NK cell isolation kit from Miltenyi ? thanks

how can i identify the LRR domain of a TLR?

In mucosal tissues, IgA is the most abundant. IgA is transported at high levels from the serosal side to the lumen by the pIgR. Bacterial stimulation of the intestinal epithelium induces the production of IgA. However, translocation of bacteria over the intestinal epithelium results in increased induction of IgG. IgG1 and IgG3 increase following exposure to the protozoa Falciparum. Whereas IgG4 is more dominantly present in soluble antigen responses and helps to prevent or reduces IgE-mediated allergic responses.

IgD is expressed first during B cell development, followed by IgM. Although the role of IgD is still partly illusive, recently IgD has been implicated in various innate immune functions: http://www.nature.com/ni/journal/v10/n8/abs/ni.1748.html.

The biology of the regulation of IgA1 and IgA2 is intriguing. Both are secreted equally efficiently by pIgR. However, as mentioned above, levels could vary between tissue suggesting differential regulation of their respective expression and CSR mechanisms. The innate factor APRIL seems to advantage the production of IgA2 over IgA1, which correlates with increased expression of IgA2 in the bacteria-rich colon: http://www.sciencedirect.com/science/article/pii/S1074761307002877.

There are numerous reviews out there, but one last comment about membrane-bound versus secreted Ig: These versions both occur for all isotype classes and are independent of CSR, but do depend on alternative splicing, where the first heavy chain domain is required for membrane-bound Ig (also referred to is BCR) and the cleaved form allows for secretion of Igs as predominantly observed in (terminally) differentiated plasma blasts and plasma cells.

You might consider to convert your anti-NKG2A antibody of choice (and the respective isotype control) into Fab/F(ab)2-fragments - there are several commercial kits for that purpose that are easy to use and reliable. By using blocking Fab/F(ab)2-fragments you circumvent the potential problem of Fc-receptor-mediated activation. Good luck!

we have hands-on experience on measure  the amount of endotoxin in mouse serum

we used the following kit.it was good

Mouse Lipopolysaccharides(LPS） ELISA Kit cat.NO: CSB-E13066m

supplied by Genxbio Health Sciences Pvt. Ltd.

Yes the RNA based interference is existed in human. the newly article publish recently have uncover a group of family existed in eukaryotes and human doing RNA interference . you can find the articles under the title "DNA-guided genome editing using the Natronobacterium gregoryi Argonaute". this technique is more powerful than CRISPER-Cas9, just their plasmid has been not introduce yet. 

In order to optimize the dose of antigen you could titrate your antigen and quantify a cytokine like IFNg or IL2 using an ELISA kit. You could access, in paralel, cell proliferation.

For pyroptosis you should canonically look for activation of caspase-1 by western blot (the p10 or p20 fragments; use the antibodies SC-514 [mouse] or SC-515 [human] from Santa Cruz Biotechnology or Casper-1 from Adipogen). You may also look for the perinuclear inflammasome foci usually associated with pyroptosis that can be detected by immunofluorescence using antibodies or with the non-antibody YVAD-FLICA tag. An example can be found in this recently published paper: 

Generally, the immune response after trauma, infection, sepsis, starts first with the activation of Th1 immunity and proinflammatory cytokine production (SIRS). As our body knows, how dangereous for our  organs can an uncontrolled hyperinflammation be (multiorgan dysfunction syndrom, multiorgan failure, failure of microcirculation, death due to cytokine storm even in the absence of microorganisms), approximately 10-24 hours later starts with the compensatory antiinflammatory response (CARS)) -  with the production of Th2 (IL-4, ...) cytokines and production of antiinflammatory cytokines (IL-10). The  aim of this is to perform a balance. If this compensatory antagonistic response is too strong, the Th1 immunity together with inflammation is strongly inhibited, and it can lead to immune depression even immune paralysis that might be the cause of development of e.g. polymicrobial sepsis. Note: the higher of hyperinflammation is, the higher the compensatory antiinflammatory response will be.

Cave: Patients with head trauma or patients suffering from immune deficiency can from the beginning start with Th2 immunity and CARS, that means without hyperinflammation and Th1 immunity actovation.

Hi Gaurav,

Thanks for the answer. In fact, I was aware of this system (a sort of hemodialysis) to remove endotoxin by polymyxin.  Rather I was curious to know about the fate of endotoxin trapped by its binder in vivo. If we consider reticuloendothelial system, it is likely to invoke innate immune system in somewhat altered manner (LPS in the endosomal compartment also detected by innate PRRs, until and unless it is degraded by the enzymatic system therein).

The best answer is "probably" yes. Your readout will be the deciding factor. If you are looking for:

Then yes, it would probably work. The following paper showed that licensing of the inflammasome is rapid and only requires on the order of 5 minutes to engage:

Effectively, the inflammasome requires post-translational licensing. The long duration priming that most people use is usually to synthesize IL-1b and additional amounts of components such as NLRP3. However, NLRP3 is present in most cells at a low level constitutively and IL-18 is definitely constitutive. As such, a brief priming with LPS or some other "signal 1" stimulus will be sufficient for NLRP3 inflammasome priming.

Using them together would probably have the intended effect, but you run the risk of the P2X7 engagement with ATP throwing the system off of balance since it is detected and responded to within a few 10s of milliseconds. It's best to do a one-two punch with a brief 5-minute priming phase followed by ATP.

Dear Younes,

Most haemocytes in your figure are granulocytes (arrows in the Fig. that I am attaching). In those granulocytes the central area appears filled with acidophilic granules, which hide the nucleus, and the peripheral area is hyaline. Although you do not mention the procedure, I guess you have deposited a drop of haemolymph onto the slide and allowed the haemocytes to adhere spontaneously onto the slide for a while; by this way, granulocytes spread on the slide surface and adhere onto the slide, thus resulting in this appearance with a peripheral hyaline area (ectosplasm) and a central area (endoplasm) filled with granules. There are also some smaller haemocytes (double arrow in the Fig. I am attaching), which do not show almost any cytoplasm, with a patent nucleus; this smaller haemocytes could be hyalinocytes or undifferentiated stem-like cells. With the procedure of spontaneous adhesion onto the slide, hyalinocytes and stem cells are mostly lost because they have much lower ability to spread and adhere than granulocytes; this is because those smaller haemocytes show reduced or almost null cytoplasm.

If you want to keep all the cell types in the slide you could either cytocentrifuge haemolymph or to use slides covered with an adhesive film; slides covered with poly-L-Lysine are frequently used. You can contact me for further explanation or any other information on this issue. I am also attaching a paper on mussel haemocyte morphology. Please, confirm that you have read this message. Best wishes,

Antonio

just because they are close does not make them the same!  The endometrium is a completely different environment and I would say not influenced by what is going on in the cervical region 

This work might help!

www.ncbi.nlm.nih.gov › NCBI › Literature › PubMed Central (PMC)

by YK Lieu - ‎2012 

Impaired adult myeloid progenitor CMP and GMP cell function in conditional .... No abnormality was detected in the pIpC-treated mybf/+/MxCre mice (data not shown). ... Cell surface expression of CD11b, CD41 (a marker of megakaryocytic ... in the CD11b+Gr-1- compartment also expressed elevated surface CD115 and ...

Hi Elise, sorry for the late reply! I'm happy to hear that! I haven't been working on CLEC-1 anymore for a long time. CLEC-1 was part of my PhD project and I was looking for a ligand but couldn't find one. I'm still very curious about it though...

We've published our CLEC-1 data in 2012 (Scand J Immunol. 2012 Mar;75(3):282-92. The human C-type lectin-like receptor CLEC-1 is upregulated by TGF-β and primarily localized in the endoplasmic membrane compartment.) We didn't find a ligand. However our approach was not very systematic, instead we tried whatever we could get our hands on for free. This is what we say in the paper: 'Potential candidates tested were known ligands of other members of the myeloid subfamily of C-type lectin-like receptors (e.g. zymosan, oxLDL and podoplanin), intracellular pathogens (e.g. Toxoplasma gondii and Listeria monocytogenes), ligands of intracellular Toll-like receptors (e.g. the TLR9 ligand CpG) and several primary cell types and cell lines.'

I wish you all the best with your CLEC-1 project!

@Namita, thank you very much for your answer.

It's worth pointing out in case you didn't realise that Hergen Spits is the foremost authority on human ILC. To add to his ST2 comment: I've only tried the MBL anti-human ST2 (clone 2A5) and it was nothing like I'd seen with the mouse. I would very much like to know what a good anti-human ST2 clone is.

There's no single reason why macrophages or other myeloid-derived cells autofluoresce, but some of the varied reasons may include the presence and concentration of NAD(P)H, flavins, porphyrin, or lipofuscin.

Personally, I've noticed that it gets worse when macrophages are primed/activated with LPS.

Here is a paper that discusses more: http://www.jleukbio.org/content/88/3/597.full

You can make F(ab')2 fragments of your antibodies - there are lots of protocols for this available, but the most common way to do it is add pepsin for a few hours to cleave the Fc portion of the antibody. This will still block the receptor, but without the Fc portion it won't activate complement or Fc-receptor-mediated responses. This assumes that your antibody is a blocking antibody, and if you are not sure of this then you should test it in vitro first.

I don't find this surprising, the human NK cells will simply contain a high amount of perforin and granzyme B to begin with.  The activation status of NK cells may be a better indicator of actual cytolytic capacity at a given time if perforin/granzyme B levels are similar.  Think of NK cells as a loaded gun, but pulling the trigger requires the right balance between activating and inhibitory signals.

CD107a expression is commonly used to indicate degranulation and therefore should correspond to cytolytic capacity (since this relates to release of lytic granules such as perforin and granzyme B).  If anything, the protein amount of perforin and granzyme B per cell may slightly increase following activation or stimulation.

Dear Thomas,

Thanks for asking this very important question.  You are absolutely right.   PMN are the primary mediators of the rapid host defense against most bacterial and fungal pathogens.  Cancer chemotherapy or reactions to cytotoxic drugs are known to cause iatrogenic neutropenias leading to severe immune deficiency.  Activation of neutrophils and to enable their phagocytic properties requires increased oxygen consumption of molecular oxygen.  The consumed oxygen is then utilized by NADPH oxidase generating ROS.  Some of the suitable methods to quantify ROS generation are as follows:

1.  Chemilunminescence amplified by luminol or isoluminol

2.  the absorbance change following reduction of cytochrome c

3.  The fluorescence increase upon oxidation of PHPA (p-Hydroxyphenyacetate).

I will be happy to provide you with the details of these techniques should you be interested.

Best Wishes,

Omer

How long is the ATP added for and how are you preparing your ATP solutions? What organism is your LPS from? You can do a western blot for intracellular pro-IL1b induction to determine if your priming step is the issue. I do 1 ug/mL for 4 hours and get robust induction as determined by both WB and IF.

I would recommend at least 30-60 minutes of ATP exposure. You can use a membrane-impermeant dye like ethidium bromide to determine if the P2X7 receptors are being activated by the ATP. I would recommend doing immunofluorescence for ASC or Caspase-1 or YVAD-FLICA to see if you have perinuclear specks that are indicative of inflammasomes.

5 mM ATP for 30-60 minutes will kill your cells, for sure. You can use a caspase-1 inhibitor like YVAD-CHO or YVAD-CMK and if you see a reduction in death then you are good to go.

Westerns on concentrated supernatants for IL1b or Caspase-1 are also a sure-sign, or ELISA for IL1b.

You have a mixed extract with potentially 100s of biologically active compounds.  Thus, this is almost to be expected.  Further purification is indicated. 

Hi Noha,

for transfection RAW macrophages I use a new transfection reagent called Viromer.

Its a synthetic polymer zero in charge. Due to an active endosome escape mechanism I get higher transfection efficiency compared to standard reagents.

Please find attached some exemplary data.

Feel free to contact me in case of question: Olivia.Zabel@lipocalyx.de

Best, Olivia

Hi Srinivas, the only way to get reliable and robust data from neutrophils is to analyse them almost immediately after the sample has been taken.  

At 6 hours ~30-50% of the neutrophils in a sample will be dead, and by 24 hours, 70-90% will be dead (dead=apoptotic).  The best advice I can give you is that you should stain your cells as soon as you are able, and then analyse them immediately after that.  There is nothing available that will preserve neutrophils in an unaltered state for more than a couple of hours (significant phenotypic changes occur in whole blood neutrophils incubated for even 4 hours ex vivo), so you will not be able to obtain data that are relevant to the in vivo situation by delaying your sample processing and analysis.

In serum the immunoglobulin G and complement proteins  can improve the phagocytosis of bactéria.

Hi,

Pig monocytes do represent a quit big group of cells which are characterized by different types of surface markers including CD14. Subgroups of monocytes including antigen-presenting cells exert different functions. I am wondering about the purpose of your cell isolation ?

Have a nice weekend !

Regards,

andrea

I believe you can find useful information in references

Antibody dependent cellular cytotoxicity ADCC                                                    www.youtube.com/watch?v=bqD-bwJemlY                                                                        Antibody-dependent cellular cytotoxicity (ADC) is a tripartite immune response whereby cytophilic antibody (Ab) directs NK cells to lyse sensitized targets. In this scheme, Ab may bind to Fc receptors on NK cells which then recognize targets specific to the Fab portions of such Ab; alternatively, cytophilic Ab bound to targets are secondarily recognized and lysed by NK cells. ADC has been demonstrated to be a potent antiviral cytotoxic mechanism both in vitro and in vivo against common human viruses including herpes simplex (HSV), measles, Epstein-Bar (EBV), and influenza virus. In addition, ADC responses have been demonstrated to play a role in the control of retroviral infections and retrovirus-induced tumors in animal models.                                                                                       Opsonization: Opsonization is the process by which a foreign particle, particularly a microbe, is coated with plasma proteins (opsonins) so as to facilitate the attachment and internalization of that particle by a professional phagocytic cell. In general, the process refers to coating of the microbe with immunoglobulin molecules (antibodies) that are specific for antigenic determinants on that organism, or with 3 complement proteins (particularly C3b) deposited on the surface of the organism via either the classical or alternative activation pathways.

The presence of these plasma proteins on the surface of the microbe facilitates their sequential interaction with 3 immunoglobulin receptors (Fc receptors) or complement receptors (CR) on the phagocyte surface. These interactions result in encirclement of the particle by the cytoplasmic membrane of the phagocytic cell, until the particle is contained within a membrane-bound vacuole (phagosome) within the cell.

Dear Evan,

We tested this using 0.1 mg 1A8 (BioXCell) i. p. (Day 0). We saw an approx. 80% depletion which lasted until Day 5. Neutrophil numbers started to increase on Day 6 and returned to normal on Day 7. Higher doses of 1A8 did not cause much more severe depletion in our hands but may last a few days longer (we did not test it). You should be careful about how to stain for neutrophils in the presence of depleting doses of 1A8 since the large circulating 1A8 concentration blocks the binding of additional (e. g. fluorescently labeled) 1A8 molecules to the epitope. We used an Ly6B (7/4) antibody to label neutrophils and gated out monocytes based on FSc/SSc. The 7/4 mAb works fine even if neutrophils are saturated with 1A8. You may also consider using the NIMP-R14 mAb for neutrophil depletion. It gave us much more robust (practically complete) depletion which lasted for 3-5 days. The specificity also seemed to be OK. You can look up Weber et al., J Exp Med 2015 (in press; link below) for further details.

Best wishes,

Attila

Depends on what kind of inhibitor you want to use.

There is a number of mAbs that inhibit either the ligand-binding domain of CR3, such as CBRM1/2 or bind to the beta I domain and block the relay of allostery (TS1/18 for example).

If you are looking for a small molecule inhibitor, Roche has developed some. Namely XVA143. And I think Genentech has 2 similar compounds.

Otherwise, CR3 binding is metal-ion dependent, so if you just add EDTA, you should abolish its function. 

Hope this helps.

A Google search on "Bovine lactoferrin antioxidant pdf returned about 2,14,000 hits

Antioxidant Effects of Bovine Lactoferrin on Dexamethasone ...

first page dump:

by L Safaeian - ‎2014 - ‎Related articles

Nov 19, 2013 - International Scholarly Research Notices is a peer-reviewed, open access journal covering a wide range of subjects in science, technology, ...

Missing: search

Effect of selenium-saturated bovine lactoferrin (Se-bLF) - DRO

by H Burrow - ‎2011 - ‎Cited by 6 - ‎Related articles

Apr 5, 2012 - bovine lactoferrin ... The states of all antioxidant enzymes (glutathione peroxidase (GPx), glutathione reductase (GR), ... Search Google Scholar.

Antioxidant enzyme activities of iron-saturated bovine ... - DRO

by H Burrow - ‎2011 - ‎Cited by 17 - ‎Related articles

Dec 5, 2011 - Antioxidant enzyme activities of iron-saturated bovine lactoferrin (Fe-bLf) in human gut epithelial cells under ... Search Google Scholar.

Patent CA2141961C - Antioxidant - Google Patents

The present invention provides a safe antioxidant applicable for food, drug medicine, non-medical drug and other products. ... Advanced Patent Search .... Kogyo Campany), bovine lactoferrin (manufactured by Sigma Company), hydrolysate of ...

Patent US7326775 - purification of transferrins ... - Google

Feb 5, 2008 - Advanced Patent Search ... 3) purifying said antioxidant-treated lactoferrin with at least one polyphenol to form purified lactoferrin; and .... Oral administration of bovine LF (40 mg/day) in healthy human volunteers (n=17) ...

lactoferrin 339615-76-8 - The Good Scents Company

Food Additive : Functional use(s) - antioxidants. ... Scientific Opinion on bovine lactoferrin:page or pdf ... Google Scholar : Search, Google Books : Search.

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Dec 12, 2014 - Your ads will be inserted here by. Google Adsense. .... antioxidant activity of an oral supplementation of bovine lactoferrin in humans. Using an

Lactoferrin - Advanced Health and Life Extension

Lactoferrin boosts immune function and has antioxidant properties. ... Advanced Health & Life Extension. Google. Custom Search ... The lactoferrin concentration in bovine (cows) milk is only 0.5% to 1.0% while human breast milk can contain ...

Full Text - Poultry Science - Oxford Journals

by L Wang - ‎2008 - ‎Cited by 24 - ‎Related articles

Institution: Google Indexer; Sign In as Personal Subscriber ..... (2008) found the similar effect of bovine lactoferrin, an antioxidant, can improve antioxidant ...

Patent US4668771 - Method for separating bovine ... - Google

Advanced Patent Search ... Method for separating bovine lactoferrin from cow's milk and purifying same ... A method as claimed in claim 1 wherein the bovine lactoferrin adsorbed to the ..... US7326775, 2 Dec 2005, 5 Feb 2008, En-N-Tech, Inc. purification of transferrins using surfactants, antioxidants and flavonoids, than ...

Interleukin-12 (IL-12), an important immuno-modulator for cell-mediated immunity, shows

significant potential as a vaccine adjuvant and anti-cancer therapeutic.Different cell types and soluble factors are involved in the induction of IFN-γ. Thus, in addition to IFN-γ-producing cells (T cells, NK cells, and macrophages) and accessory cells (macrophages and dendritic cells) a number of cytokines are known to participate directly or indirectly in the induction of IFN-γ. Among the soluble factors, IL-12 is of special interest, being capable of directly inducing IFN-γ in T and NK cells and promoting T cell differentiation in the IFN-γ-producing Th1 subset. Splenocyte suspensions is prepared from spleens of 6- to 8-wk-old mice by pressing spleens through a wire grid. Pooled cells from three or four animals are  suspended in serum-free DMEM, adjust to a concentration of 107/ml, and place (2 × 106/well) in 96-well plates (Nunc, Roskilde, Denmark). They are then cultured in the presence or the absence of stimulating agents (10 μl/well) at 37°C in a humidified atmosphere containing 8% CO2 for 24 h. Culture supernatants for determination of IFN-γ are stored in aliquots at −80°C until use.IFN-γ in supernatants of splenocyte cultures and in murine plasma is estimated by  ELISA. The limit of IFN-γ detection is 60 pg/ml. IL-12 (p70) in supernatants of macrophage cultures is estimated by ELISA using anti-IL-12 mAb (C17.8) and biotin-labeled anti-IL-12 mAb (C15.6; PharMingen, Hamburg, Germany) as described previously (45). The limit of detection is15 pg/ml.

RIG-I-like receptors (RLRs)  cytoplasmic sensors of pathogen-associated molecular patterns (PAMPs) within viral RNA. play a major role in pathogen sensing of RNA virus infection to initiate and modulate antiviral immunity.

The RLRs detect viral RNA ligands or processed self RNA in the cytoplasm to triggers innate immunity and inflammation and to impart gene expression that serves to control infection.RLRs cooperate in signaling crosstalk networks with Toll-like receptors and other factors to impart innate immunity and to modulate the adaptive immune response

patients with single nucleotide polymorphisms (SNPs) in genes(RIG-I, MDA5, and LGP2 )encoding become highly susceptible to RNA virus infection  

There are numerous studies to demonstrate the antiviral response that is induced by poly I:C in mice (following an injection). It is through induction of type I interferons (IFNs), which through induction of the IFN-activatable genes exert the antiviral effects. Please see papers beginning 1975.

I wonder whether Jonathan really  had asked us  about which orпan/tissue from one can best get macrophages for culturing (as most of answerers  evidently understood him) or about organs in which a significant population of resident (sic!) macrophages plays  an important functional role. I believe these are two different questions. and answers are not the same.  It is rather easy to get macrophages for in vitro experiments from peritoneum by washing it (even without preliminary activation) but I am not sure that  their role in situ is great¹⁾. On the contrary, the red pulp of the spleen would not be able to catch senescent erythrocytes from the blood for utilisation if it were not rich in resident macropgages, but if you look simply for a convenient source of macropgahes for culture,  the spleen or the liver (in which Kupfer cells also have a very important function) would not be the best choice.

====

¹⁾ Moreover, I am not sure that peritoneal macrophages meet definition of the resident ones as a tissue-specific poulation of macrophages self-maintaining locally  with only  minimal contribution from circulating monocytes.

Many thanks Kelli for all advices and papers.

I need a pure population of epithelial cells, not organoids, for co-culture model with fibroblasts. So I would like to grow them around 2 weeks.

Btw. Congratulation of really good paper.

Supraphysiological levels of E2 can provoke anorexia, so keep an eye on food intake and weight loss.

I suggest measure IL-p70 and not p40.

Good luck.

Hallo,

First of all it depends on the standards used. The WHO (Fa. Behring) has produced 50000 liters of pure standard CRP. If this standard is used, then the first problem is solved. Then, there is the reproducibility of different labs. If you take the Ringversuche of DGKL for glucose, for example, which is so well defined almost as no other molecule!. The target value of one was 100 mg/100 ml. The values offered by well known german laboratiries varied between 80 and 180 mg/dl. Now you have the answer!

greetings

Sighart Golf

Yes, indeed. It is quite often very difficult to prove a hypothesis or to find some more hints to a theory and against the paradigms.

It took decades to find some experimental support for some parts of Einstein´s theories, or, more recently, for the Higgs boson.

And there often are more than just one, two or three people involved in great findings. In the 1990s an immunologist in Zürich got the Nobel prize, but I am sure many expected Hugh McDevitt to get it as well since his findings were earlier. A few years later, McDevitt got a different award - and I think it was Irv Weissman who mentioned in the laudatio in a Montreal meeting that this was the "biggest award in immunology"  (it was much more money than a Nobel prize). Not sure what the former Nobel winner sitting behind me thought at that time, but it is not the recipient of an award who makes the decisions.

So, it may just remain hard to know and understand all the details which lead to the decisions of the Nobel committee.

Sorry,I meant 

Dear Rune,

 

Dear Daxing

Please review http://www.jimmunol.org/content/162/3/1582.full.pdf

Hakeem Sam and Mary M. Stevenson have described specific antibodies that may resolve your issues.